Circadian-Dependent Intermittent Fasting Influences Ischemic Tolerance and Dendritic Spine Remodeling.

BACKGROUND: Preconditioning by intermittent fasting is linked to improved cognition and motor function, and enhanced recovery after stroke. Although the duration of fasting was shown to elicit different levels of neuroprotection after ischemic stroke, the impact of time of fasting with respect to the circadian cycles remains unexplored. METHODS: Cohorts of mice were subjected to a daily 16-hour fast, either during the dark phase (active-phase intermittent fasting) or the light phase (inactive-phase intermittent fasting) or were fed ad libitum. Following a 6-week dietary regimen, mice were subjected to transient focal cerebral ischemia and underwent behavioral functional assessment. Brain samples were collected for RNA sequencing and histopathologic analyses. RESULTS: Active-phase intermittent fasting cohort exhibited better poststroke motor and cognitive recovery as well as reduced infarction, in contrast to inactive-phase intermittent fasting cohort, when compared with ad libitum cohort. In addition, protection of dendritic spine density/morphology and increased expression of postsynaptic density protein-95 were observed in the active-phase intermittent fasting. CONCLUSIONS: These findings indicate that the time of daily fasting is an important factor in inducing ischemic tolerance by intermittent fasting.

An antioxidant and anti-ER stress combination therapy elevates phosphorylation of α-Syn at serine 129 and alleviates post-TBI PD-like pathology in a sex-specific manner in mice.

Clinical studies have shown that traumatic brain injury (TBI) increases the onset of Parkinson's disease (PD) in later life by >50%. Oxidative stress, endoplasmic reticulum (ER) stress, and inflammation are the major drivers of both TBI and PD pathologies. We presently evaluated if curtailing oxidative stress and ER stress concomitantly using a combination of apocynin and tert-butylhydroquinone and salubrinal during the acute stage after TBI in mice reduces the severity of late-onset PD-like pathology. The effect of multiple low doses of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) on post-TBI neurodegeneration was also evaluated. The combo therapy elevated the level of phosphorylation at serine 129 (pS129) of α-Syn in the pericontusional cortex of male mice at 72 h post-TBI. Motor and cognitive deficits induced by TBI lasted at least 3 months and the combo therapy curtailed these deficits in both sexes. At 3 months post-TBI, male mice given combo therapy exhibited significantly lesser α-Syn aggregates in the SN and higher TH+ cells in the SNpc, compared to vehicle control. However, the aggregate number was not significantly different between groups of female mice. Moreover, TBI-induced loss of TH+ cells was negligible in female mice irrespective of treatment. The MPTP treatment aggravated PD-like pathology in male mice but had a negligible effect on the loss of TH+ cells in female mice. Thus, the present study indicates that mitigation of TBI-induced oxidative stress and ER stress at the acute stage could potentially reduce the risk of post-TBI PD-like pathology at least in male mice, plausibly by elevating pS129-α-Syn level.

Key to the North American tribes and genera of herb, rose, bramble, and inquiline gall wasps (Hymenoptera, Cynipoidea, Cynipidaesensu lato).

Robust keys exist for the family-level groups of Cynipoidea. However, for most regions of the world, keys to genera are not available. To address this gap as it applies to North America, a fully illustrated key is provided to facilitate identification of the tribes and genera of rose gall, herb gall, and inquiline gall wasps known from the region. For each taxon covered, a preliminary diagnosis and an updated overview of taxonomy, biology, distribution, and natural history are provided.

Non-pharmacological interventions for traumatic brain injury.

Heterogeneity and variability of symptoms due to the type, site, age, sex, and severity of injury make each case of traumatic brain injury (TBI) unique. Considering this, a universal treatment strategy may not be fruitful in managing outcomes after TBI. Most of the pharmacological therapies for TBI aim at modifying a particular pathway or molecular process in the sequelae of secondary injury rather than a holistic approach. On the other hand, non-pharmacological interventions such as hypothermia, hyperbaric oxygen, preconditioning with dietary adaptations, exercise, environmental enrichment, deep brain stimulation, decompressive craniectomy, probiotic use, gene therapy, music therapy, and stem cell therapy can promote healing by modulating multiple neuroprotective mechanisms. In this review, we discussed the major non-pharmacological interventions that are being tested in animal models of TBI as well as in clinical trials. We evaluated the functional outcomes of various interventions with an emphasis on the links between molecular mechanisms and outcomes after TBI.

Post-stroke brain can be protected by modulating the lncRNA FosDT.

We previously showed that knockdown or deletion of Fos downstream transcript (FosDT; a stroke-induced brain-specific long noncoding RNA) is neuroprotective. We presently tested the therapeutic potential of FosDT siRNA in rodents subjected to transient middle cerebral artery occlusion (MCAO) using the Stroke Treatment Academic Industry Roundtable criteria, including sex, age, species, and comorbidity. FosDT siRNA (IV) given at 30 min of reperfusion significantly improved motor function recovery (rotarod test, beam walk test, and adhesive removal test) and reduced infarct size in adult and aged spontaneously hypertensive rats of both sexes. FosDT siRNA administered in a delayed fashion (3.5 h of reperfusion following 1 h transient MCAO) also significantly improved motor function recovery and decreased infarct volume. Furthermore, FosDT siRNA enhanced post-stroke functional recovery in normal and diabetic mice. Mechanistically, FosDT triggered post-ischemic neuronal damage via the transcription factor REST as REST siRNA mitigated the enhanced functional outcome in FosDT-/- rats. Additionally, NF-κB regulated FosDT expression as NF-κB inhibitor BAY 11-7082 significantly decreased post-ischemic FosDT induction. Thus, FosDT is a promising target with a favorable therapeutic window to mitigate secondary brain damage and facilitate recovery after stroke regardless of sex, age, species, and comorbidity.

Post-stroke depression: epigenetic and epitranscriptomic modifications and their interplay with gut microbiota.

Epigenetic and epitranscriptomic modifications that regulate physiological processes of an organism at the DNA and RNA levels, respectively, are novel therapeutic candidates for various neurological diseases. Gut microbiota and its metabolites are known to modulate DNA methylation and histone modifications (epigenetics), as well as RNA methylation especially N6-methyladenosine (epitranscriptomics). As gut microbiota as well as these modifications are highly dynamic across the lifespan of an organism, they are implicated in the pathogenesis of stroke and depression. The lack of specific therapeutic interventions for managing post-stroke depression emphasizes the need to identify novel molecular targets. This review highlights the interaction between the gut microbiota and epigenetic/epitranscriptomic pathways and their interplay in modulating candidate genes that are involved in post-stroke depression. This review further focuses on the three candidates, including brain-derived neurotrophic factor, ten-eleven translocation family proteins, and fat mass and obesity-associated protein based on their prevalence and pathoetiologic role in post-stroke depression.

Cerebroprotective Role of N6-Methyladenosine Demethylase FTO (Fat Mass and Obesity-Associated Protein) After Experimental Stroke.

BACKGROUND: FTO (fat mass and obesity-associated protein) demethylates N6-methyladenosine (m6A), which is a critical epitranscriptomic regulator of neuronal function. We previously reported that ischemic stroke induces m6A hypermethylation with a simultaneous decrease in FTO expression in neurons. Currently, we evaluated the functional significance of restoring FTO with an adeno-associated virus 9, and thus reducing m6A methylation in poststroke brain damage. METHODS: Adult male and female C57BL/6J mice were injected with FTO adeno-associated virus 9 (intracerebral) at 21 days prior to inducing transient middle cerebral artery occlusion. Poststroke brain damage (infarction, atrophy, and white matter integrity) and neurobehavioral deficits (motor function, cognition, depression, and anxiety-like behaviors) were evaluated between days 1 and 28 of reperfusion. RESULTS: FTO overexpression significantly decreased the poststroke m6A hypermethylation. More importantly, exogenous FTO substantially decreased poststroke gray and white matter damage and improved motor function recovery, cognition, and depression-like behavior in both sexes. CONCLUSIONS: These results demonstrate that FTO-dependent m6A demethylation minimizes long-term sequelae of stroke independent of sex.

An antioxidant and anti-ER stress combo therapy decreases inflammation, secondary brain damage and promotes neurological recovery following traumatic brain injury in mice.

The complex pathophysiology of post-traumatic brain damage might need a polypharmacological strategy with a combination of drugs that target multiple, synergistic mechanisms. We currently tested a combination of apocynin (curtails formation of reactive oxygen species; ROS), tert-butylhydroquinone (promotes disposal of ROS), and salubrinal (prevents endoplasmic reticulum stress) following a moderate traumatic brain injury (TBI) induced by controlled cortical impact in adult mice. Adult mice of both sexes treated with the above tri-combo showed alleviated motor and cognitive deficits, attenuated secondary lesion volume, and decreased oxidative DNA damage. Concomitantly, tri-combo treatment regulated post-TBI inflammatory response by decreasing the infiltration of T cells and neutrophils and activation of microglia in both sexes. Interestingly, sexual dimorphism was seen in the case of TBI-induced microgliosis and infiltration of macrophages in the tri-combo treated mice. Moreover, the tri-combo treatment prevented TBI-induced white matter volume loss in both sexes. The beneficial effects of tri-combo treatment were long-lasting and were also seen in aged mice. Thus, the present study supports the tri-combo treatment to curtail oxidative stress and endoplasmic reticulum stress concomitantly as a therapeutic strategy to improve TBI outcomes.SIGNIFICANCE STATEMENTOf the several mechanisms that contribute to TBI pathophysiology, oxidative stress, endoplasmic reticulum (ER) stress, and inflammation play a major role. The present study shows the therapeutic potential of a combination of apocynin, tert-butylhydroquinone, and salubrinal to prevent oxidative stress and ER stress and the interrelated inflammatory response in mice subjected to TBI. The beneficial effects of the tri-combo include alleviation of TBI-induced motor and cognitive deficits and lesion volume. The neuroprotective effects of the tri-combo are also linked to its ability to prevent TBI-induced white matter damage. Importantly, neuroprotection by the tri-combo treatment was observed to be not dependent on sex or age. Our data demonstrate that a polypharmacological strategy is efficacious after TBI.

Antioxidant therapies in traumatic brain injury.

Oxidative stress plays a crucial role in traumatic brain injury (TBI) pathogenesis. Reactive oxygen species (ROS) and reactive nitrogen species (RNS) formed in excess after TBI synergistically contribute to secondary brain damage together with lipid peroxidation products (reactive aldehydes) and inflammatory mediators. Furthermore, oxidative stress, endoplasmic reticulum stress and inflammation potentiate each other. Following TBI, excessive oxidative stress overloads the endogenous cellular antioxidant system leading to cell death. To combat oxidative stress, several antioxidant therapies were tested in preclinical animal models of TBI. These include free radical scavengers, activators of antioxidant systems, Inhibitors of free radical generating enzymes and antioxidant enzymes. Many of these therapies showed promising outcomes including reduced edema, blood-brain barrier (BBB) protection, smaller contusion volume, and less inflammation. In addition, many antioxidant therapies also promoted better sensory, motor, and cognitive functional recovery after TBI. Overall, preventing oxidative stress is a viable therapeutic option to minimize the secondary damage and to improve the quality of life after TBI.

The Arthropod Associates of 155 North American Cynipid Oak Galls.

The identities of most arthropod associates of cynipid-induced oak galls in the western Palearctic are generally known. However, a comprehensive accounting of associates has been performed for only a small number of the galls induced by the estimated 700 species of cynipid gall wasps in the Nearctic. This gap in knowledge stymies many potential studies of diversity, coevolution, and community ecology, for which oak gall systems are otherwise ideal models. We report rearing records of insects and other arthropods from more than 527,306 individual galls representing 201 different oak gall types collected from 32 oak tree species in North America. Of the 201 gall types collected, 155 produced one or more arthropods. A total of 151,075 arthropods were found in association with these 155 gall types, and of these 61,044 (40.4%) were gall wasps while 90,031 (59.6%) were other arthropods. We identified all arthropods to superfamily, family, or, where possible, to genus. We provide raw numbers and summaries of collections, alongside notes on natural history, ecology, and previously published associations for each taxon. For eight common gall-associated genera (Synergus, Ceroptres, Euceroptres, Ormyrus, Torymus, Eurytoma, Sycophila, and Euderus), we also connect rearing records to gall wasp phylogeny, geography, and ecology -including host tree and gall location (host organ), and their co-occurrence with other insect genera. Though the diversity of gall wasps and the large size of these communities is such that many Nearctic oak gall-associated insects still remain undescribed, this large collection and identification effort should facilitate the testing of new and varied ecological and evolutionary hypotheses in Nearctic oak galls.

DNA damage and repair following traumatic brain injury.

Traumatic brain injury (TBI) is known to promote significant DNA damage irrespective of age, sex, and species. Chemical as well as structural DNA modification start within minutes and persist for days after TBI. Although several DNA repair pathways are induced following TBI, the simultaneous downregulation of some of the genes and proteins of these pathways leads to an aberrant overall DNA repair process. In many instances, DNA damages escape even the most robust repair mechanisms, especially when the repair process becomes overwhelmed or becomes inefficient by severe or repeated injuries. The persisting DNA damage and/or lack of DNA repair contributes to long-term functional deficits. In this review, we discuss the mechanisms of TBI-induced DNA damage and repair. We further discussed the putative experimental therapies that target the members of the DNA repair process for improved outcome following TBI.

Postischemic supplementation of folic acid improves neuronal survival and regeneration in vitro.

Supplementation of folic acid (FA) is beneficial to several neurological diseases because it promotes notch signaling and neurogenesis and reduces blood homocysteine levels. We hypothesized that postischemic supplementation of FA is beneficial for neuronal survival and regeneration. The objective of the present study was to determine the postischemic neuroprotective and neuroregenerative efficacy of FA supplementation and its effects on various cellular processes in vitro. This work benefited from the use of FA and glucose-free media to better assess the ischemic neuroprotection provided by FA supplementation. The postischemic supplementation of FA significantly improved cell viability, and the improvement was primarily by obstructing the oxygen-glucose deprivation (OGD)-activated apoptosis. Furthermore, postischemic treatment with FA significantly reduced the mitochondrial membrane depolarization and the formation of acidic organelles triggered by OGD. Moreover, FA's effect on neuroregeneration following OGD was evaluated by measuring the cell proliferation and neurite outgrowth length. Treatment with FA enhanced cell proliferation and neurite outgrowth significantly. Thus, these results revealed some of the mechanisms by which FA supplementation provided neuroprotection and neuroregeneration following ischemic injury and highlighted the need for further research into the potential of folic acid as a clinical drug for ischemic stroke.

Folic Acid Exerts Post-Ischemic Neuroprotection In Vitro Through HIF-1α Stabilization.

The constant failure of single-target drug therapies for ischemic stroke necessitates the development of novel pleiotropic pharmacological treatment approaches, to effectively combat the aftermath of this devastating disorder. The major objective of our study involves a multi-target drug repurposing strategy to stabilize hypoxia-inducible factor-1 α (HIF-1α) via a structure-based screening approach to simultaneously inhibit its regulatory proteins, PHD2, FIH, and pVHL. Out of 1424 Food and Drug Administration (FDA)-approved drugs that were screened, folic acid (FA) emerged as the top hit and its binding potential to PHD2, FIH, and pVHL was further verified by re-docking, molecular dynamics (MD) simulation and by Drug Affinity Responsive Target Stability (DARTS) assay. HIF-1α stabilization by FA was demonstrated by the nuclear translocation and increased green fluorescence emission of HIF-1α using HIF1α-GFPSpark tag vector. Further, FA treatment enhanced the cell survival following oxygen glucose deprivation and its neuroprotective mechanism was elucidated by measuring the expression of BAX, NFE2L2, VEGF, and EPO genes in a time-dependent manner (5 and 11 h following FA treatment). VEGF and EPO expressions were significantly increased by 5.41- and 1.35-folds, respectively, whereas BAX expression reduced by 4-fold at 11 h post-FA treatment. NFE2L2 expression was elevated (1.65-fold) at 5 h with no major difference at 11 h post-FA treatment. The chicken chorioallantoic membrane (CAM) assay demonstrated the pro-angiogenic potential of FA as evidenced by an increased blood vessel density and branching. The present study elucidates for the first time that the post-ischemic neuroprotection exerted by FA may be attributed to its HIF-1α stabilization and pro-angiogenic properties.

Hypoxia Mimetic Agents for Ischemic Stroke.

Every year stroke claims more than 6 million lives worldwide. The majority of them are ischemic stroke. Small molecule-based therapeutics for ischemic stroke has attracted a lot of attention, but none has been shown to be clinically useful so far. Hypoxia-inducible factor-1 (HIF-1) plays a crucial role in the transcriptional adaptation of cells to hypoxia. Small molecule-based hypoxia-mimetic agents either stabilize HIF-1α via HIF-prolyl hydroxylases (PHDs) inhibition or through other mechanisms. In both the cases, these agents have been shown to confer ischemic neuroprotection in vitro and in vivo. The agents which act via PHD inhibition are mainly classified into iron chelators, iron competitors, and 2 oxoglutarate (2OG) analogs. This review discusses HIF structure and key players in the HIF-1 degradation pathway as well as the genes, proteins and chemical molecules that are connected to HIF-1 and how they affect cell survival following ischemic injury. Furthermore, this review gives a summary of studies that used PHD inhibitors and other HIF-1α stabilizers as hypoxia-mimetic agents for the treatment of ischemic injury.

Systematic review and stratified meta-analysis of the efficacy of carnosine in animal models of ischemic stroke.

Carnosine is a naturally occurring pleotropic dipeptide which influences multiple deleterious mechanisms that are activated during stroke. Numerous published studies have reported that carnosine has robust efficacy in ischemic stroke models. To further evaluate these data, we have conducted a systematic review and meta-analysis of published studies. We included publications describing in vivo models of ischemic stroke where the neuroprotective efficacy of carnosine was being evaluated through the reporting of infarct volume and/or neurological score as outcomes. Overall efficacy was evaluated using weighted mean difference random effects meta-analysis. We also evaluated for study quality and publication bias. We identified eight publications that met our inclusion criteria describing a total of 29 comparisons and 454 animals. Overall methodological quality of studies was moderate (median = 4/9). Carnosine reduced infarct volume by 29.4% (95% confidence interval (CI), 24.0% to 34.9%; 29 comparisons). A clear dose-response effect was observed, and efficacy was reduced when carnosine was administered more than 6 h after ischemia. Our findings suggest that carnosine administered before or after the onset of ischemia exhibits robust efficacy in experimental ischemic stroke. However, the methodological quality of some of the studies was low and testing occurred only in healthy young male animals.

Novel RIPK3 inhibitors discovered through a structure-based approach exert post-ischemic neuroprotection.

Necroptosis or programmed necrosis is evident in various neurological disorders such as ischemic stroke. Receptor interacting serine/threonine protein kinase 3 (RIPK3) is one of the crucial targets of necroptosis and inhibition of this protein exerts neuroprotection. However, knowledge regarding the three-dimensional structure and binding site information of this protein is lacking. In the present study, structure-based in silico methods were implemented to identify the key amino acids in the RIPK3 binding site that might be responsible for ligand interactions. Further, novel RIPK3 inhibitors were identified through a dual ensemble screening strategy. Three inhibitors exhibited binding to RIPK3 in micromolar concentrations and exerted post-ischemic neuroprotection in vitro.